Mounting evidence suggests that deregulation of protein phosphorylation pathways is a primary contributor to the development and progression of Alzheimer's disease (AD) and other aging-related neurodegenerative diseases. In particular, sustained activation of extraceilular signal-regulated kinase (ERK), a major subtype of the mitogen-activated protein kinase (MAPK) family of sedne/threonine kinases, occurs very ready in AD, is implicated in neurofibdllary tangle formation, and, moreover, leads to neuronal degeneration. It follows that understanding the biochemical bases for the regulation of ERK signaling will lead to a greater understanding of the neurodegenerative process. Activation of ERK requires phosphorylation of both the threonine and tyrosine residues in an activation loop TXY sequence by upstream ERK/MAPK dual-specificity kinases (MEKs). Thus, inactivation of ERK requires dephosphorylation of only one of the phosphoamino acids, although the mechanisms by which inactivation occurs in various tissues and under different conditions remains to be fully characterized. We have established recently that the major constitutively-expressed ERK phosphatase in rat brain cerebral cortex is a microsomal membrane-enriched, okadaic acid-sensitive, PP2A-like enzyme. Moreover, we have found in preliminary studies that the constitutive ERK phosphatase is inactivated by the thiol-reactive nitrogen oxide species (RNOS) peroxynitrite (ONOO-) and nitroxyl (HNO). These findings are consistent with the hypothesis that the ERK phosphatase contains an essential reactive cysteine residue(s) that renders the enzyme sensitive to inactivation by RNOS and, perhaps, reversible nhibition by glutathione-mediated S-thiolation. Increases in both RNOS generation and protein S-thiolation are relevant to conditions of neurodegeneration. Excessive inhibition of constitutive ERK phosphatase activity by either of these pathways would be expected to promote sustained activation of ERK and deregulate ERK signaling. The research proposed herein will employ biochemical methods to fully characterize the protein identity, phosphoamino acid specificity, bases for inhibition by RNOS, and postulated sensitivity to S-thiolation of the identified PP2A-iike constitutive ERK phosphatase.